Characterisation of human fetal mesenchymal stem cells

Mesenchymal stem cells (MSCs) are present in various tissues of fetal and adult origin. I isolated and expanded MSCs from human fetal 1st trimester livers and observed that they have a shorter cell doubling time compared to adult bone marrow-derived MSCs. Fetal MSCs differentiated into osteogenic, chondrogenic and adipogenic lineages when induced in vitro. Furthermore, functional assays of adipocyte-differentiated fetal and adult MSCs demonstrated that the intracellular pathways, expression of proteins involved in lipolysis, lipolytic activity and secretion of leptin and adiponectin were similar to that of mature adipocytes, although some differences were noted.

Fetal MSCs did not differentiate into adipocytes as readily as adult MSCs did. Fetal MSCs exhibit a similar morphology and surface expression pattern as adult MSCs. Analysis of surface proteins by flow cytometry showed presence of CD29, CD44, CD73, CD105 and CD166 but not of the hematopoietic markers CD34, CD45 or CD14. HLA class I expression was lower in fetal than in adult MSCs. HLA class II could not be detected on the surface, and fetal MSCs had no intracellular deposits of HLA class II as did adult MSCs. Seven days of IFNgamma exposure was required for full surface expression of HLA class II, compared to two days for adult MSCs. Fetal MSCs, like adult MSCs, did not upregulate HLA class II during differentiation to osteogenic, chondrogenic and adipogenic lineages.

Further analysis of differences between fetal and adult MSCs was conducted by gene array. Transcriptomes of fetal and adult MSCs differed mainly in genes coding for developmental, cell cycle regulatory and immunologic transcripts. Fetal MSCs showed increased expression of transcripts involved in germ plasm and limb patterning and in brain and early muscle development. The expression of transcripts implicated in cell cycle promotion, chromatin regulation and DNA repair were also more abundant in fetal MSCs. Transcripts with reduced expression in comparison to adult MSCs were those involved in smooth muscle and keratinocyte differentiation. Fetal MSCs expressed fewer transcripts for immunologic genes, which could imply that fetal MSCs are less immunologically mature than adult MSCs.

The effect of fetal MSCs on proliferating lymphocytes was investigated by co-culture experiments and mitogen assays. Mitogen stimulation of lymphocytes was inhibited by fetal MSCs. Neither undifferentiated nor differentiated fetal MSC induced proliferation of allogeneic lymphocytes. Unlike adult MSCs, fetal MSCs did not inhibit proliferating lymphocytes, while fetal MSCs treated with IFNgamma for seven days did. These results suggest that fetal MSCs are immunologically privileged cells and have potentials for allogeneic transplantation.

MSCs may be used in cellular therapies. A female fetus with multiple intrauterine fractures, diagnosed as severe osteogenesis imperfecta, was transplanted with HLA-mismatched male fetal MSCs in the 32nd week of gestation. At 35 weeks, the baby girl was delivered by cesarean section. At nine months of age a centromeric XY-specific probe revealed 0.3% of XY-positive cells in a bone marrow biopsy. Whole Y genome FISH staining showed a median of 7.4% Y-positive cells. Patient lymphocyte proliferation against donor MSCs was not observed in co-culture experiments performed in vitro before and after MSC injection, indicating that the patient was not immunised against the allogeneic cells. During the first two years of life three fractures were noted and growth followed the same curve. Thus, allogeneic mis-matched MSCs can be safely transplanted in utero to a patient with severe OI, where the cells engraft in bone. To conclude, fetal MSCs may be a valuable source for transplantation.

List of scientific papers

I. Gotherstrom C, Ringden O, Westgren M, Tammik C, Le Blanc K (2003). Immunomodulatory effects of human foetal liver-derived mesenchymal stem cells. Bone Marrow Transplant. 32(3): 265-72.
https://pubmed.ncbi.nlm.nih.gov/12858197

II. Gotherstrom C, Ringden O, Tammik C, Zetterberg E, Westgren M, Le Blanc K (2004). Immunologic properties of human fetal mesenchymal stem cells. Am J Obstet Gynecol. 190(1): 239-45.
https://pubmed.ncbi.nlm.nih.gov/14749666

III. Ryden M, Dicker A, Gotherstrom C, Astrom G, Tammik C, Arner P, Le Blanc K (2003). Functional characterization of human mesenchymal stem cell-derived adipocytes. Biochem Biophys Res Commun. 311(2): 391-7.
https://pubmed.ncbi.nlm.nih.gov/14592427

IV. Gotherstrom C, West A, Liden J, Lahesmaa R, Le Blanc K (2004). Difference in gene expression between human fetal and adult mesenchymal stem cells. [Manuscript]

V. Le Blanc K, Gotherstrom C, Ringden O, Hassan M, McMahon R, Horwitz E, Anneren G, Axelsson O, Nunn J, Ewald U, Norden-Lindeberg S, Jansson M, Dalton A, Astrom E, Westgren M (2004). Mesenchymal stem cells engraftment in bone following in utero transplantation in a patient with severe osteogenesis imperfecta. [Manuscript]